Project Summary The Cancer Genome Atlas has uncovered mutations in human ovarian cancer (OVCA) genomes that potentially drive tumorigenesis and alter cell metabolism to meet the crucial requirements of tumor cells. Whereas mutations in metabolic enzymes hardwire metabolism to tumorigenesis, they are relatively infrequent in OVCA. More often, cancer metabolism is altered by the abundance and activity of the metabolic enzymes through ubiquitin-proteasome proteolysis. We applied bioinformatic tools and meta-analysis to analyze genes in the ubiquitin-proteasome proteolysis and determined their molecular interactions with cell metabolism. The most significantly modulated gene identified from our analyses is ubiquitin specific peptidase 13 (USP13). The proposed studies are based on the three novel findings: 1) In-depth analysis of OVCA genomes identified copy number gains of USP13 gene in 29.3% (158 out of 538) of high- grade serous OVCA, but only in 3.7% of breast cancer and in 0% of colorectal cancer, suggesting that USP13 amplification is a unique and frequent genomic event in OVCA; 2) Two potential deubiquitination targets of USP13, ATP citrate lyase (ACLY) and oxoglutarate dehydrogenase (OGDH), are key regulators that determine glutaminolysis, tricarboxylic acid (TCA) cycle and lipid synthesis; 3) Metabolism of reactive stromal cells in tumor microenvironment (TME) is reprogrammed through an elevated glutamine anabolic pathway, which confers atypical metabolic flexibility and adaptive mechanisms in stromal cells, allowing them to harness carbon and nitrogen from noncanonical sources to synthesize glutamine in nutrient-deprived conditions in ovarian TME. Despite the fact that many ovarian tumors show increased uptake of glucose and glutamine and elevated lipid metabolism, no molecular mechanisms have been identified and little progress has been made towards harnessing the potential therapeutic use of these observations. The bottleneck is to find key genomic alterations that drives OVCA cell metabolism. Here, we propose that 1) USP13 amplification drives ovarian cancer cell metabolism by upregulating ACLY and OGDH, and 2) A synthetic lethal approach to target PIK3CA and USP13 in in OVCA harboring the USP13 amplicon for desirable therapeutic outcomes. We will test these hypotheses in three aims: Aim 1: To determine the USP13 induced metabolic alterations in OVCA with USP13 amplification; Aim 2: To determine molecular mechanisms for the regulation of USP13 activity; Aim 3: To assess the effects of USP13 inhibition in vivo using human ovarian tumor models. The ability of USP13 in rewiring metabolism and its druggability fuel the interest in targeting USP13 and cancer metabolism in OVCA, leading to the development of new therapeutics.